PROJECT 2: MUSCLE CELL TRANSPLANTATION IN CANINE DMD Muscle-derived cell transplantation has the potential to effectively treat many human diseases, including muscular dystrophy. Studies in mdx mice demonstrate that normal muscle-derived cells engraft into skeletal muscle, effectively restore dystrophin expression and reconstitute the satellite cell pool. However, immune rejection of donor cells prevented long-term engraftment in human trials. We have induced immune tolerance in cxmd canines using a clinically relevant regimen of bone marrow transplantation, and show donor muscle-derived cell engraftment and survival for at least 24 weeks in the absence of immunosuppression. Our broad and long-term objective is to identify a cell population and accessory factors required for efficient engraftment of myogenic stem cells into diseased canine skeletal muscle that can be translated to human trials in muscular dystrophies. In addition to the immune tolerant cxmd model, we have developed a canine-to-mouse xenotransplantation model to rapidly and quantitatively compare canine muscle cell engraftment. We will use both of these models to test the broad hypothesis that transplantation of donor muscle-derived cells will successfully reconstitute normal muscle to clinically significant levels in a canine model of Duchenne muscular dystrophy. Demonstrating successful engraftment in a random-bred, large animal model will provide the basis for establishing a new clinical regimen of treating human skeletal muscle diseases. The Specific Aims of our proposal will (Aim 1) determine whether using specific subpopulations of muscle-derived cells and modulating signaling pathways will enhance engraftment of transplanted donor cells and reconstitution of the satellite cell compartment, and (Aim2) determine whether the muscle stem cell transplantation techniques that are successful in mice can be directly translated to the immune tolerant canine cxmd model of Duchene muscular dystrophy. RELEVANCE (Seeinstructions): Together, these studies will optimize cell isolation and delivery techniques using pre-clinical therapeutic cell transplant studies in mice and canines. The significance of this study is that canine models of hematopoietic cell transplantation and solid organ transplantation have been demonstrated to accurately predict clinical outcomes in humans. These studies will establish the best protocols for muscle cell transplantation and lead to future human clinical trials of muscle cell transplants for the treatment muscular dystrophies and for the delivery of systemic factors.